The WT1 gene was isolated from human chromosome 11p13 as one of the genes etiologically associated with Wilms' tumor on the basis of an analysis of WAGR syndrome involving such complications as Wilms' tumor, aniridia, urogenital anomalies, mental retardation, etc. It is known that its expression is substantially confined to the fetal kidney, spleen, testis, and ovary [Molecular Medicine, Vol. 32, No. 5, p. 502 (1995)].
The inventors of the present invention reported previously that the level of WT1 gene expression is high in acute leukemia, that this level of expression is inversely correlated with prognosis, and that the so-called MRD (minimal residual disease) in acute leukemia can be detected by determining said expression level [Blood, Vol. 84, No. 9, p. 3071 (1994)].
The expression level of the WT1 gene can be determined by the well-known reverse transcribed polymerase chain reaction (RT-PCR; Kawasaki, E. S. et al., Amplification of RNA, in PCR Protocol, A Guide to Methods and Applications, Academic Press, Inc., San Diego, 21-27 (1991)) and this determination has proved clinically significant as far as leukemia is concerned.
On the other hand, a solid cancer is a neoplastic condition in which cancer cells grow in a solid mass and is quite distinct from leukemia in which tumor cells severally grow, infiltrate, and metastasize.
Heretofore, much research has been undertaken into solid cancers but no clinical marker corresponding to said WT1 gene in leukemia has been reported as yet.
Meanwhile, myelodysplastic syndrome (MDS) is an acquired hematopoietic disturbance featuring preleukemia and a refractory decrease in peripheral blood cell count. Its definite diagnosis depends on detection of a morphological abnormality of blood cells and, here, disease typing has so far been made according to the FAB (French-American-British Group) classification.
It is known that MDS mentioned above progresses to acute leukemia at a high rate and the incidence of leukemic transformation is said to be not less than 40% in the high-risk group and as high as 10-15% even in the low-risk group.
With regard to the prognosis and prognostic factors for MDS, the above-mentioned FAB classification is well known and the risks for acute leukemia are said to be high blast ratios of bone marrow and peripheral blood, clusters of blasts on bone marrow biopsy, and complicated chromosomal aberrations [Naika Gaku (Internal Medicine), p. 1711, Sep. 25, 1995, Asakura Shoten].
As diseases which may undergo leukemic transformation in the similar way, myelofibrosis, polycythemia vera, aplastic anemia, primary thrombocytosis, and paroxysmal nocturnal hemoglobinuria, among others, are known but there has not been available an established technology for detecting MDS and other diseases progressing to leukemia, i.e. an atypia with morphological features distinct from those of the normal tissue or cells.
Meanwhile, in patients with leukemia, a therapeutic modality which comprises performing an allogenic bone marrow transplantation (alloBMT), i.e. transfusion of hematopoietic stem cells, for avoiding irreversible hematopoietic disorders and concurrently instituting an anticancer drug therapy, a radiation therapy, and/or the like for exterminating leukemic cells in cooperation with the immune reactions of the patient and thereby achieving a complete cure has been practiced with some success. More recently, peripheral blood stem cell transplantation (PBSCT) or autologous BSCT (ABSCT), in lieu of autologous bone marrow transplantation (ABMT) which is a kind of said bone marrow transplantation, has been adopted by clinicians with increasing avidity because of its numerous advantages.
In such a bone marrow or peripheral blood stem cell transplantation, the myelocytes or peripheral blood stem cells from a donor with a substantially or completely compatible HLA antigen are transplanted into the patient. The hematopoietic stem cells in the graft then start normal hematopoiesis enlisting the help of the interstitial cells and hematopoietic factors in the bone marrow, thus leading to a recovery of hematopoiesis. While such a bone marrow or peripheral blood stem cell transplantation is rewarded with a fair success, the practice involves the grave problem of recurrence after transplantation in addition to the scarcity of donors and the graft-related complications.
To overcome the above-mentioned recurrence and other problems, it is most essential to insure that the very tissue or cells to be transplanted are free of cancer cells such as leukemic cells, that is to say the safety of the donor's bone marrow or peripheral blood stem cells, but there has not been available an established technology for testing whether leukemic cells are included among myelocytes or peripheral blood stem cells.
It is, therefore, an object of the present invention to provide a technology for quantitating tumor cells which is of value for the diagnosis of solid cancers, more particularly a quantitative test utilizing a clinical marker correlated with solid cancers.
Another object of the present invention is to provide a technology for detecting atypia utilizing a novel marker by which the risk of acute leukemic transformation, for instance, can be predicted with greater accuracy taking into consideration the course and prognosis of MDS in particular.
It is a still another object of the present invention to provide a novel technology which permits detection of leukemic cells in the tissue for use in said bone marrow transplantation or peripheral blood stem cell transplantation.